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BackgroundOlfactory ecto-mesenchymal stem cells (OE-MSC) are mesenchymal stem cells derived from the lamina propria of the nasal mucosa. They display neurogenic and immunomodulatory properties and were shown to induce recovery in animal models of spinal cord trauma, hearing loss, Parkinsons’s disease, amnesia, and peripheral nerve injury. As a step toward clinical practice, we sought to (i) devise a culture protocol that meets the requirements set by human health agencies and (ii) assess the efficacy of stem cells on neuron differentiation.MethodsNasal olfactory mucosa biopsies from three donors were used to design and validate the good manufacturing process for purifying stem cells. All processes and procedures were performed by expert staff from the cell therapy laboratory of the public hospital of Marseille (AP-HM), according to aseptic handling manipulations. Premises, materials and air were kept clean at all times to avoid cross-contamination, accidents, or even fatalities. Purified stem cells were cultivated for 24 or 48 h and conditioned media were collected before being added to the culture medium of the neuroblastoma cell line Neuro2a.ResultsCompared to the explant culture-based protocol, enzymatic digestion provides higher cell numbers more rapidly and is less prone to contamination. The use of platelet lysate in place of fetal calf serum is effective in promoting higher cell proliferation (the percentage of CFU-F progenitors is 15.5%), with the optimal percentage of platelet lysate being 10%. Cultured OE-MSCs do not show chromosomal rearrangement and, as expected, express the usual phenotypic markers of mesenchymal stem cells. When incorporated in standard culture medium, the conditioned medium of purified OE-MSCs promotes cell differentiation of Neuro2a neuroblastoma cells.ConclusionWe developed a safer and more efficient manufacturing process for clinical grade olfactory stem cells. With this protocol, human OE-MSCs will soon be used in a Phase I clinical based on their autologous transplantation in digital nerves with a neglected injury. However, further studies are required to unveil the underlying mechanisms of action.

International audience; Background: Platelet-rich plasma improves engraftment after fat transfer. However, the effects of platelet dose have never been investigated. The authors used magnetic resonance imaging to compare surviving graft volumes in mice after administration of four different formulations (microfat alone, and three platelet-rich plasma-enriched microfat mixes). Methods: The authors used a random, double-blinded, fat transfer protocol using three different platelet levels: 1 million (low-dose), 500 million (medium-dose), and 1000 million (high-dose) platelets/ml, and fat alone (control). The authors grafted 0.4 ml of the 70/30 platelet-rich plasma-enriched microfat mixtures (0.4 million, 200 million, and 400 million platelets per 0.12 ml for the low-dose, medium-dose, and high-dose mixtures, respectively) or 0.4 ml of microfat alone into 22 nude mice and monitored surviving graft volumes every month for 3 months. Then, the authors histologically analyzed all grafts to assess neoangiogenesis status and fat integrity. Results: Three-dimensional magnetic resonance imaging showed that the median surviving graft volumes at 3 months were 9.5 percent (interquartile range, 0 to 25 percent; p = 0.003) (high-dose), 4.1 percent (interquartile range, 0 to 18 percent; p = 0.001) (medium-dose), and 18 percent (interquartile range, 8 to 38 percent; p = 0.41) (low-dose) compared to 36 percent (interquartile range, 28 to 53 percent) for the control value. The histologic integrity of microfat-alone grafts was significantly better than those of the other grafts, although the high-dose and low-dose grafts exhibited higher levels of neoangiogenesis. Conclusion: Higher platelet levels in microfat grafts were associated with poor graft survival in nude mice; a clinical review would be appropriate.